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High Specific Activity [Samarium-153] EDTA for Imaging of Experimental Tumor Models
John W. Tse, Leonard I. Wiebe and Antoine A. Noujaim
J Nucl Med. 1989;30:202-208.
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(Print ISSN: 0161-5505, Online ISSN: 2159-662X)
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High SpecificActivity [Samarium.-153]EDTA
for Imaging of Experimental Tumor Models
John W. Tse, Leonard I. Wiebe, and Antoine A. Noujaim
Faculty ofPharmacy and Pharmaceutical Sciences, University ofAlberta, Edmonton, Canada
Enrichedsamariumoxide(98.2% 1@Sm2O3)
was irradiatedin low neutronflux and high
neutronflux reactorsto produce1@Smwith specificactivitiesof 14.3 GBqand22.1TBq
mmor'1Sm,respectively,at the timeof use.Formulationof 1@Smas [1@SmJEDTA,
with a
1:10 molar ratio of SM:EDTA, provided a stable radiotracer in vitro and in vivo. High specific
activity [153Sm]EDTAshowed superior uptake in cell culture (20.8 ±0.9% vs. 5.5 ±0.1% for
6 and 600 pmol Sm per 106cells, respectively) and better tumor index @values
(51 vs. 37 at
10.9 nmol and 1.09 @moI
Sm kg1 , respectively) in the BDF1mouse/Lewis lung tumor model.
High specific activity [1@'Sm]EDTAscintigrams of Copenhagen x Fisher rats bearing
transplantedDunningR3327 tumorsclearlydelineatedthe tumorswithin6 hr, with moderate
liverand boneuptakeand low soft-tissuebackground.The injectedradiotracerunderwent
rapidcentralcompartmentclearanceandwhole-bodyelimination.Theabsenceof observed
adversehistopathologicaltoxicitycombineswith highimagequalitywithin6 hr, to supportthe
clinicaltumor-imaging
potentialof thisagent.Comparative
studieswith[67Gajcitrate
at molar
equivalentdosesindicatedthat highspecificactivity[1@Sm]EDTA
was a superiorradiotracer
in these in vitro and in vivo models.
J Nucl Med 30:202—208,1989
he known tumor affinity of Group III elements
such as gallium and indium has stimulated interest in
the use of radiolanthanides and radioactinides for both
radiotherapy and scintigraphy. Gallium-67 (67Ga) and
indium-i 11 (‘
‘
‘In)have found broad application in
diagnostic nuclear medicine (1,2). The use of radiolan
thanides, including samarium-i53 (‘53Sm)
for tumor
imaging was pioneered two decades ago (3). Although
O'Mara et al. (3) proposed that the radiolanthanides
would be useful bone scanning agents, ‘“Sm
has re
cently received attention as a therapeutic radiophar
maceutical
for the palliative
treatment
of metastatic
bone cancer(4—6).In spite ofthe complex decay scheme
which includes various beta (Em,@,0.8 MeV) and elec
that can be obtained in high yield and with high specific
activity from low-cost, enriched ‘52Sm2O3.
Radioactive
decay yields a principal gamma photon of 103 keV
(29.8%), with little (0.6%) high-energy gamma emission
and a half-life of 46.27 hr (7,16). Quantitative tumor
uptake and scintigraphy ofhigh specific activity [‘53Sm]
ethylenetnaminetetraacetic
acid (EDTA) in experimen
tal tumor models are now reported, in comparison with
a lower specific activity formulation and with [67Ga]
citrate.
METhODS AND MATERIALS
Production of ‘@Sm
tron (to 0. 1 MeV) radiations ( 7) associated with ‘“Sm Low specific activity [‘53Smjsamarium
oxide (1.1 mCi mg―
decay,
this reactor-produced
radiolanthanide
emits
gamma radiations which are well-suited for high effi
ciency, high resolution gamma cameras. Favorable tu
mor uptake has been demonstrated for several radiolan
thanides and a number of ‘“Sm
tumor uptake studies
have been reported (8—15).
Samarium- 153 is a reactor-produced radiolanthanide
Received May 16, 1988;revision accepted Sept. 21, 1988.
For reprints contact: Leonard I. Wiebe, Faculty of Pharmacy
and Pharmaceutical Sciences, University of Alberta, Edmonton,
Canada T6G2NB.
202
Tse,Wiebe,
andNoujaim
Sm203; 14.3 GBq.mmol―)was produced by thermal neutron
irradiation (4 hr at 1 x 1012n .cm'2.@')
ofenriched [‘52Sm]
Sm203(Oak RidgeNational Laboratory;98.2% 152Sm)at the
University of Alberta SLOWPOKE Reactor Facility. High
specific activity ‘“Sm
was produced by irradiation (7 days at
2 x lO'4n•cm'2
.scc') of the enriched Sm203 at the Atomic
Energy of Canada Ltd. research reactor facility (AECL Com
mercial Products Division, Chalk River, Canada). The specific
activity at time of use was estimated to be 1.7 Ci mg'' Sm203
(22.1 TBq mmor' Sm). Yields and radionuclidic purity were
similar to those reported by Ehrhardt et al (1 7). Radionuclidic
purity was determined by high resolution Ge(Li) gamma
TheJournalof NuclearMedicine
Downloaded from jnm.snmjournals.org by on April 20, 2014. For personal use only.
spectrometry. The 70- and l03-keV photopeaks of ‘“Sm
were
the main gamma emissions detected, along with low-energy
photons arising from trace amounts of unidentified radionu
clidic contaminants. Radionuclidic purity was estimated to be
> 99%, based on gamma
counts
at the time of use.
Preparation of t'53SmIEDTA
Irradiated [‘53Sm]Sm2O3
was dissolved in sufficient lN HQ
to produce a solution of 1 mg SmCl3 ml―.This was added to
an aqueous solution of disodium ethylenediaminetetracetic
acid (Na2EDTA) so that the molar ratio was Sm:EDTA =
1:10. After standing for 30 mm, the pH was adjusted to 6.5
by slow, careful addition of iN NaOH, afterwhich the solution
was filtered through a 0.22-tim filter (Miffipore Ltd., Missis
sauga, Canada). These solutions were diluted with distilled
water to a final activity of 370 kBq in 0.1—0.2ml.
Samarium-l53 carbon-l4 (‘4C)
EDTA was prepared by
mixing the [‘53Sm]SmCl3
solution (37 MBq) with a solution
of['4CIEDTA in 0.iN HG (370 kBq;4 GBq mmol'')(Amer
sham Canada Ltd., Oakville, Canada). After incubation for
30 mm, an aqueous solution ofNa2EDTA was added to make
a solution with a molar ratio Sm:EDTA = 1:10. After an
additional 30 mm the pH was adjusted to 6.5 with iN NaOH
and filtered through a 0.22-tim filter. Stability of this formu
lation, as judged by colloid formation in RPM! growth me
dium, was determined by ultracentnfugation as reported else
where (14).
Gallium-67 citrate (Merck Frosst Inc., Montreal, Canada)
was obtained as a sterile, non-pyrogenic solution containing 9
ng Ga ml―with a concentration of 2 mCi (74 MBq) mr'.
Carrier Ga citrate was added for in vitro studies so that uptake
could be compared with Sm uptake at identical molar concen
trations.
Radiochromatography and Radiometry
Radiochemical purity of the radiotracer formulation was
determined using ascending paper chromatography. Strips of
Whatman No. 1 paper (25 x 130 mm) were run in pyri
cline:ethanol:water = 1:2:4 for 2 hr, after which they were
dried, cut into transverse strips (10 mm) and counted in a
well-type gamma counter (Smith Kline Beckman, Irvine, CA;
Model 8000). Cellulose gel thin layer chromatography (TLC)
(Terochem Laboratories,Edmonton, Canada; Eastman Ko
dak 6064 plates) were also run with this solvent system. After
development, the chromatograms were cut into 10-mm strips
and counted in a gamma counter. In the case of dual-label
(‘53Sm/'4C)
studies, the strips were re-counted after 2 1 days
by adding distilled water (1.5 ml) to wet the strip, letting it
stand overnight and then mixing with scintillation fluor
in RPMI-l640 medium which contained heat-inactivated fetal
calf serum (FCS 15%) and glutamine (0.29 mg m1―).The
EMT-6 cells were maintained in vitro as a monolayer in
Waymouth's medium supplemented with FCS (10%) and
glutamine (0.29 mg m1―).Suspensions of the respective cell
lines for in vitro studies were prepared by washing monolayer
cells with Versene buffer, followed by a brief incubation (1
mm) with 0.1% trypsin (Gibco, Grand Island, NY). Cells were
then washed (2 x 10 ml) with the respective growth medium,
followed by dilution to 1 x 106 cells ml'' in that medium.
Cell viability was determined by the trypan blue exclusion test
as described elsewhere (15). Viability normally exceeded 98%.
In in vitro uptake studies, cell suspension aliquots (1 ml; 1 x
106cells) were incubated with the radiotracer formulation (0.6
nmol radionuclide for Melanoma 2AB cells and 0.006 nmol
radionuclide for EMT-6 cells) at 37°for predetermined inter
vals. After incubation, cells were pelleted by centrifugation
and washed twice with normal saline, and then both the cell
pellets and the combined supernatants were radioassayed.
Experiments were run in triplicate, with corrections for non
viable cells.
In vivo biodistributionof ‘@Sm
and 67Garadiotracers
Whole-body tissue distribution studies were carried out on
male BDF, mice (28—30g) bearing Lewis lung carcinoma
which had been implanted in the flank. Radiotracer solutions
(370 kBq in 0.1—0.2ml) were slowly injected via the tail vein
at mass doses of 1.09 @mol
and 10.9 nmol kg' (for low and
high specific activity formulations, respectively) into mice
which had tumors with diameters of 5—7.5
mm.
At specified time intervals, animals were killed; an aliquot
ofblood and the entire lungs, liver, spleen, kidney, femur and
tumor were collected, weighed and radioassayed using a well
type gamma scintillation counter. The remaining carcass was
counted in a large-volume well counter (Picker International,
Northford, CT; Model Autowell II). In mice dosed with
[‘53Sm]['4CJEDTA,
tissue specimensweighing 60 mg were
taken for gamma analysis, then stored for 40 days (for ‘53Sm
decay) prior to radioassay for ‘4C.
These samples were solu
bilized in a mixture of 2 ml Protosol (Amersham) in 100 @il
water. After dissolution, samples were decolorized by the
addition of a few drops of hydrogen peroxide (30% v/v),
neutralized by addition of glacial acetic acid (0.1 ml) and
mixed with scintillation fluor (DuPont Company) (15 ml).
Stannous chloride (three drops of a 4% w/v solution in 0. iN
HO) was added to reduce chemiluminescence, and samples
were dark-adapted at 5°for 30 mm prior to counting in a
liquid scintillation counter. Counting efficiency corrections
were based on the H-number external standard (Smith Kline
(DuPont Company,Inc., Montreal,Canada;AquasolII)prior Beckman), with interpolation from a set of quenched stand
to beta counting in a liquid scintillation counter (Smith Kline ards prepared in an identical manner but containing a known
amount of [‘4C]hexadecane
(Amersham).
Beckman, Irvine, CA; Model 9000). The stability of [‘“Sm]
Whole-body scintigraphy of male Copenhagen x Fisher
EDTA was determined in the presence of[45Ca]calciumchlo
g) bearing subcutaneously-implanted Dunning
ride (Terochem) (1.63 MBq @tg@).
The solutions were mixed, rats (300—325
allowed to stand for 30 mm and then applied to thin layer R-3327H prostatic tumors was undertaken using a gamma
chromatographic (TLC) plates and analyzed as described camera(Siemens Medical Systems, Des Plaines, IL) fitted with
a high resolution collimator designed for technetium-99m.
above.
Scintigrams of not less than 100,000 counts were recorded
after i.v. injection (femoral vein) of either 1.09 @molor 10.9
In vitro cell uptake studies
Human Melanoma 2AB and murine EMT-6 cell lines were nmol Sm per kg body weight (3.7 MBq) for low and high
obtained from the Cross Cancer Institute, Edmonton. The specific activity Sm, respectively. In a preliminary study, a
Melanoma 2AB cells were maintained in vitro as monolayers formulation with Sm:EDTA = 1:2 was also used with a Sm
Volume30 •
Number2 •
February1989
203
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dose of 1.09 @molper kg body weight. In some studies, rats
were housed individually in metabolic cages after dosing, with
food and water ad libitum. Urine was collected at various
intervals for further analysis. In all cases, quantitative whole
body counting was undertaken using the gamma camera, with
the rat held in fixed geometry during measurements.
Limited studies on weight gain and histopathologic effect
ofi.v. doses ofSm-EDTA (0. 1, 1.09 and 10.9 @imol
kg') were
undertaken using male ALAS mice (20—23g). Weights were
recorded on Days 1, 4, 12, 18, 24, 28, 32, 36, and 48, and
histopathologic examinations were performed at Days I, 20,
48, and 64. The male offspring ofpregnant female ALAS mice
dosed with i.v. [SmJEDTA (I .09 @molkg'') were killed 48
and 64 days after birth. Organs examined included lungs,
liver, kidney, and femur. Preliminary radiodosimetry esti
mates were made using the MIRD method (18).
a 2-hr period. Samarium uptake was 3.6 ±0.5 and 5.3
±0.3% per 106 cells at 1 hr and 2 hr, respectively,
compared to 3.0 ±0.2 and 3.9 ±0. 1 for [67Ga]citrate.
Both radiotracers were studied at radionuclide concen
trations ofO.6 nmol per ml per incubation tube. Uptake
studies with low specific activity [‘53Sm]['4C]EDTA
showed similar uptakes for ‘53Sm,
but ‘4C
uptakes were
always
40% of the ‘53Sm
value. Studies with high
specific activity [‘53Sm]['4C]EDTAshowed similar ‘4C
accumulations, but [‘53Sm]
values increased much more
rapidly. Data are presented in Table 1.
Quantitative whole-body distribution of ‘53Sm,
‘4C,
and 67Garadioactivity was compared after i.v. injection
of (a) low specific activity [‘53Sm]['4C]EDTA, (b) high
specific activity [‘53Sm]EDTA,and (c) [67Ga]citrate, in
BDF, mice bearing Lewis lung carcinoma. Samarium
153 and 67Ga were found to be concentrated in several
RESULTS
tissues, but in all cases, [‘4C]EDTA
accumulations were
more transient and of a lower magnitude than concen
Samarium-i53
EDTA was formulated with 1:10
Sm:EDTA molar concentration ratio to ensure com
trations of the other two radionuclides. Samarium-153
biodistribution at both specific activities was character
ized by rapid blood clearance to very low levels (<
0.01% injected dose mr') compared with 67Ga (0.8%
injected dose mr'), thereby leading to greatly elevated
tumor:blood and tissue:blood ratios for ‘53Sm
compared
plete chelation of ‘53Sm.
TLC ofthe product at pH 6.5
showed a single spot at Rf 0.9, whereas at 1:1 stoicheo
metry, two approximately
equal spots at Rf 0.1
(‘53SmCl3)
and 0.9 were present. This formulation was
stable when incubated for 1 hr in RPM! growth me
dium, with only 3.5 ±0.6% precipitable by centrifu
gation at 144,000 g for 1 hr; incubation in distilled
water resulted in 3.3 ±2.3% sedimentation in control
with 67Ga. Specific activity-related effects included a
twofold increase in % injected dose (% I.D.) g'' of tissue
(tumor, liver, kidneys, lung and spleen) for a 100-fold
experiments. All sedimentation experiments were per
formed in triplicate at a concentration
EDTA per ml per incubation tube.
of6
decrease in ‘53Sm
dose, with no appreciable change in
@@mol
[‘53Sm] bone uptake. The high specific activity 67Ga showed a
four- to sixfold increase in % I.D. g' soft tissue at 1 hr,
The stability of Sm-EDTA in the presence of ionic
but these values converged at later intervals; however,
bone uptake at low specific activity demonstrated the
calcium, which occurs in blood in appreciable concen
trations, was tested in two ways. Incubation (1 hr) of
[‘53Sm]Cl3
with an equimolar amount ofCa-EDTA (Rf
inverse effect, with 67Ga levels nearly twice as high as
those found after injection of high specific activity
[67Ga]citrate. Tissue:blood ratio maxima invariably oc
curred 48 hr after injection for ‘53Sm,whereas the
maxima always occurred at 24 hr for ‘4C
and usually at
24 hr for 67Ga. Selected data are presented in Table 2.
0.9) led to formulation of['53Sm]EDTA in > 90% yield,
whereas incubation of [‘53Sm]EDTA
with 45CaC12(Rf
0.05) resulted in no measurable formation of [45Ca]
EDTA.
Uptake of [‘53Sm]EDTAby Melanoma 2AB cells in
The influence of Sm:EDTA ratios at a Sm dose of
1.09 @tmolkg―whole-body weight was investigated in
culture increased as a function of incubation time over
TABLE 1
Uptakeof [1@―Sm][14C]EDTA
by Melanoma2AB cellsinDeterminations%
Suspension.Dataare Means±s.d. for3
Incubation
time(mm)
1@Sm214C21@―Sm/14C25
1@Sm1
14C1
3.7 ±0.2
4.3 ±0.3
1.6 ±0.1
1.9 ±0.2
4.7 ±0.4
2.1 ±0.1
2.2 ±0.2
8.2 ±0.12.2
0.645
4.9 ±0.1
5.0 ±0.1
2.2 ±0.1
2.3 ±0.1
2.2 ±0.1
2.2 ±0.1
9.1 ±0.32.4
10.5 ±2.92.6
5.6 ±0.4
2.7 ±0.0
2.1 ±0.1
14.9 ±3.12.5
5.5 ±0.1
2.8 ±0.1
2.0 ±0.1
20.8±0.93.2
0.7I
‘@‘Sm).2
Each tube
204
1@Sm/14C1
0.310
0.220
0.230
1.160
1.4120
Each
Radioactivity in Cells
tube
contained
0.6
contained
0.006
nmole
Sm,
nmole
Sm:EDTA
Sm,
Tse,Wiebe,
andNoujaim
Sm:EDTA
2.3 ±0.2
2.3 ±0.2
= 1 :1 0 (low
= of 1:10
specific
(high
2.8 ±0.71
5.5 ±0.22.2
.8 ±0.31
±0.22.5
±0.13.8
±0.33.9
±0.14.0
±0.65.9
±0.16.6
.6 ±
±
±
±
±
±
±
activity
Specific
activity
1@Sm).
The Journal of Nuclear Medicine
Downloaded from jnm.snmjournals.org by on April 20, 2014. For personal use only.
2Quantitative
TABLE
Whole-Body Distribution of High Specific Activity [1@'Sm]EDTA,Low Specific Activity [1@Sm][14C]EDTA
and [67Ga]Citratein BDF1MiceBearingTransplantedLewisLungCarcinoma.
AnimalsTime
Dataare Means±s.d. for 4
AfterInjection
hr)Blood
Tissue
Radiotracer
1
[1@Sm][14C]EDTA 0.22 ±0.06b
0.00[@Sm][@C1EDTAc
0.04[@SmJEDTAd 0.54±0.15
0.0067Ga
1.60 ±0.21
0.22@Ga
citrate
3.40±0.29
0.25Bone
citrated
24.51±12.1
[1@'Sm][14C]EDTA 12.2±2.5
142(48)[@@Sm][l4C]EDTAc
7.7(24)[@Sm]EDTA―
0.56±0.16
(24)67Ga
12.4±2.2
5(48)@Ga
citrate
20.1±1.9
2.5(48)Tumor citrated
10.7±I .2
[1@Sm]['4C]EDTA' 0.85±0.21
17(48)[@SmJ[l4C]EDTAc
6
24
48
0.04±0.01
0.08±0.04
0.37±0.08
1.89±0.25
12.2±0.21
17.1±3.21
O.13±0.08
18.6±8.1
10.3±2.89
7.25±1.65
1.23±0.31
0.02±0.00
0.05±0.02
0.02±0.01
0.81±0.05
2.84±0.77
12.9±2.41
0.36±0.29
15.9±4.9
12.9±2.7
7.43±0.77
0.74±0.09
<0.01 ±
0.43±
0.01±
0.80±
0.96±
5.02±1.23
0.18±0.06
6.98±0.71
15.8±0.65
8.77 ±2.35
0.39±0.10
Tissue:blood
Maximum±
s.d.(time,
867 ±
8.6 ±
853 ±87
20 ±
9.2 ±
65 ±
2.2(24)[@Sm]EDTAd
0.62±0.28
0.24±0.14
0.27±0.11
0.26±0.17
16(48)67Ga
1.95±0.19
1.70±0.14
1.01±0.11
0.72±0.12
72 ±
0.9(24)°@Ga
citrate
0.88±0.09
0.76±0.19
1.37±0.75
1.23±0.42
1.7±
0.3(48)Uver citrated
7.11 ±3.62
[1'@Sm]['4C]EDTA
65(48)[@Sm][l4CJEDTAc
4,88 ±1.71
5.37±1.78
6.17±2.03
3.04±0.67
5.95±0.89
2.09 ±0.52
4.15 ±0.42
2.2 ±
691 ±
5.0(24)[@Sm]EDTAd
0.72±0.18
0.28±0.16
0.43±0.26
0.22±0.12
8.7±
88(48)67Ga
12.8±0.83
2(24)@Ga
citrate
3.09±0.68
2(48)Kidney citrated
15.7±7.82
[‘@Sm][14C]EDTA' 3.19 ±1.12
53(48)[@Sm][l4C]EDTAc
10(24)[@Sm]EDTAd 4.41 ±3.09
30(48)67Ga
13.89 ±2.08
13.1±1.97
4.63±1.85
13.9±1.41
3.02 ±1.21
1.27±1.02
5.99±1.20
12.1±1.81
17.3±1.39
12.5±2.76
1.55 ±0.43
0.88±0.55
1.56±1.01
7.67±0.84
16.0±4.12
11.8±2.96
0.97 ±0.29
0.67±0.07
1.48±0.29
767 ±
21 ±
12 ±
161 ±
18 ±
148 ±
0.4(24)°1Ga
citrate
0.3(48)a
citrated
weight).b
‘@Sm distribution
s.d.C
% injected
dose
(above).d
14C
distribution
weight).•
High specific
after
4.3±
5.07±0.92
2.34±1.05
2.18±0.48
1.61±0.52
2.7±
16.4±8.22
7.08±0.71
6.25±4.01
5.30±1.32
5.5 ±
[1@Sm]['4C]EDTA
injection
(1 .09 mol Sm
kg1
body
per 9 ±
after
activity
injection
of
radiaotracer
a
(10.9
nmol
Sm or Ga kg1
body
@Ga
distribution
after[%a] citrateinjection(1.09molGakg' bodyweight).
a preliminary scintigraphic experiment with rats bear
ing implanted Dunning prostatic tumors. At a 1:2 ratio
a scintigram of comparabje quality was not seen after
injection of 67Ga. The 12-, 24-, and 48-hr scintigrams
of Sm:EDTA, early hepatic activity was evident along
provided a clear view of the tumors with virtually no
with kidney and bladder visualization,
blood or intestinal background (Fig. 1).
Whole-body retention of both low and high specific
activity [‘53Sm]EDTAand [67Ga]citrate was measured
as was the case
for the 1:10 formulation. However, the 1:2 formulation
produced a rapid and continuing intensification of ac
tivity in the liver throughout the observation period,
suggesting gradual formation of colloidal ‘53Smand
over a 48-hr period. At each specific activity, retention
of67Ga was approximately double that of ‘“Sm
(Table
trapping of the colloid in the liver. At the 1.09 @imol 3). Fecal radioactivity accounted for 1 ±0.5% and
kg' dose, the 1:10 formulation provided good deline
urine contained 80 ±4% ofthe ‘“Sm
dose after 48 hr.
ation of the skeleton, liver and the tumor within 6 hr.
High specific activity [‘53Sm]EDTA
and [67Ga]citrate
(10.9 nmol kg' whole body) were injected i.v. into the
surgically-exposed
femoral
vein
of Copenhagen
x
Fisher rats bearing transplanted Dunning R3327-AT
tumors (100 mm mean diameter). Scintigrams were
recorded at 15 and 30 mm, and 1, 6, 12, 24, and 48 hr
after dosing. Animals dosed with [‘53Sm]EDTAshowed
blood-pool and kidney images after 1 hr, but by 6 hr,
especially in the animals which had voided, tumor,
kidney, liver and bone were clearly delineated, whereas
Volume30 •
Number2 •
February1989
Clearance of ‘53Sm
from the blood occurred with an
overall half-life of < 5 mm (Table 4); preliminary
analysis ofthese data using graphical methods suggested
a complex, perhaps triexponential, clearance pattern.
Toxicity studies in mice (15 ±1 g) dosed with i.v.
Sm-EDTA (0. 1, 1.09 or 10.9 @zmolSm kg') or i.v.
normal saline, failed to reveal any significant (p < 0.05;
n = 4 per group) change in weight gain over a 64-day
observation period. During this time, the mice gained
16 ±2 g. Histopathologic examination of lung, liver,
kidney, and long bone (femur) detected no major his
205
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@
A
T
FIGURE 1
plantedDunningR3327-Hprostatic
6700
I
I'
15
tologic changes over a 64-day period, nor in the first
generation male offspring of female mice dosed at 1.09
@molSm kg' body weight. A nonsignificant number
of hyaline casts were observed in the proximal tubules
24 hr after injection ofthe two higher dose ranges, along
with a slight increase in the number of bi- and tri
nucleated regenerating liver cells.
DISCUSSION
Interest in the development of effective, low cost,
@
diagnostic radiopharmaceuticals
which have wide
spread geographic availability has led to a number of
studies of reactor-produced radionuclides, including
‘53Sm.
In the present studies, ‘53Sm2O3
was produced
in both low and high thermal neutron flux (1 x 1012
and 2 x i0'@n .cm2 . sec', respectively) research reac
tors. Yields, specific activity, and radionuclidic purity
were similar to published data (1 7,19). Preparation of
the EDTA chelate, from the ‘53SmCl3
intermediate, has
been described elsewhere for other ‘53Smcomplexes
TABLE 3
Whole-BodyRetentionof Low- and HighSpecificActivity
[1@'Sm]EDTA
and [@7Ga]
Citrateafter i.v. Injectioninto
Fisher x Copenhagen Rats Bearing Transplanted
DunningR3327-H ProstaticTumors
% of RadioactivityRemainingin Body
Timeafter
injection(hr)
6
12
24
kg1.t
. 1 .09
umol
Low SpecificActivity'
HighSpecificActivityt
1@Sm
%a
‘@Sm 61Ga
100*
98.9±0.5 95.6±0.5 99.6±0.3
18.9±2.5 83.6±1.9 47.0±0.981.6±0.9
18.8±2.174.3±3.1
42.5±1.1 75.2±2.4
19.2±2.7 59.4±0.8 36.5 ±0.9 57.2 ±0.9
19.2±4.4 44.7±0.3 23.7 ±0.5 40.5±0.4
radionuclide
IUS.,i
‘U
Whole-body scintigrams of Fisher x
Copenhagen rats bearing trans
tumors. Samarium-i 53 EDTA (upper
row) and [67Ga]citrate(lower row)
were administered at a radionuclide
dose of 10.9 nmolkg1 bodyweight.
Scintigramswere recordedat (A) 15,
30, and 60 mm, and (B) 6, 24, and
48 hr after i.v. injection.T indicates
the location of the bilaterally-im
plantedtumors.
B
IUSm
60
6
24
(4,11—15).As a note ofcaution, however, it was found
to be essential that neutralization of the [‘53Sm]
C13EDTA solution to pH 6.5 be approached slowly and
with care to avoid high local concentrations
of base
which may cause formation of hydroxides/oxides,
thereby resulting in some colloid formation. Traces of
solids could be removed by ifitration through a 0.22micron filter, without appreciable losses of radioactiv
ity.
When formulated with a tenfold molar excess of
Na2EDTA, the [‘53Sm]EDTAwas stable in both RPMI
and Waymouth growth media and in the presence of
cultured cells. At a twofold EDTA excess, chromatog
raphy indicated that
50% of the ‘53Sm
was chelated,
whereas only chelated ‘53Sm
was detected at the 1:10
ratio. Similar observations on the stability of ‘“Sm
complexes have been reported for citrates and other
ligands (13,15). The chemical nature of the [‘53Sm]
EDTA complex has not been reported, but with nine
coordination numbers, it has been pointed out that
Sm(EDTA),
Sm(EDTA)25@ and Sm2(EDTA)36 are
possible and at least two species have been detected by
TABLE4
In VivoBloodClearanceof ‘@“Sm
after i.v. Injectionof
[1@Sm]EDTA(1.09 umoleSm kg1) intoCopenhagenx
FisherRatsBearingSubcutaneously
Transplanted
DunningR-3327HProstaticTumors
•Iime
(mm)
AfterInjection
% of InjectedRadiOaCtivity
Remaining
inBIOOd
186.0±4.4532.6±
8.61018.0±1.41516.1±0.92012.6±3.0259.9±
0.9308.8
0.5455.9
0.4603.5
±
±
0.81201.9±0.8‘Mean±s.d.,n=3.
±
kg'.*
10.9 nmol radionudide
voided.•me@fl±s.d.,
not
n=3.
206
Tse,Wiebe,
andNoujaim
The Journal of Nuclear Medicine
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@
@
high performance
liquid chromatography
(HPLC)
when ‘53SmCl3
was added to EDTA in a molar ratio of
1: 10 (19). Although the 1:2 and 1: 10 formulations
produced similar scintigraphic distribution patterns in
rats at early time periods, by
6 hr after injection a
slow, absolute increase in hepatic radioactivity was
observed with the 1:2 formulation. This suggests that
colloid was gradually being formed in vivo and that
radioactivity was redistributed to the reticuloendothe
hal system without further excretion from the body.
Uptake of [‘53Sm]EDTAat low specific activities in
cell culture was quantitatively similar to uptakes of
[67Ga]citrate at similar molar concentrations.
In the
dual-labeled [‘53Sm]['4C]EDTA study, the ratio of
‘53Sm:'4C
uptake was virtually constant in human Mel
anoma 2AB cells (2.29 ±0.21 vs. 1.99 ±0.06%). At
high specific activity, there were no significant changes
in 67Ga or [‘4C]EDTAuptakes by murine EMT-6 cells,
but [‘53Sm]uptake increased by four times the low
specific activity values, to 20.7 ±0.9%. The reason for
this response may reflect differences between the mu
rine and human cells used, or simply the limited uptake
capacity which can transport or bind a larger fraction
of the smaller dose.
The biodistribution of both low or high specific ac
tivity [‘53Sm]EDTAin Lewis lung carcinoma-bearing
BDF, mice was characterized by low blood levels (1.6—
0.006% I.D. g') and low uptake by tumor (1.95—0.39%
I.D., g'), with slightly higher values for the high specific
activity formulation (Table 2). In liver and kidney, the
latter formulation produced two to three times higher
levels at all periods except at 24 and 48 hr, when specific
activity did not effect [‘53Sm]content. Carbon-l4
EDTA from doubly-labeled [‘53Sm]['4C]EDTAwas
concentrated to only 10—20%ofthe [‘53Sm]
concentra
tion in liver, which represents the stoichiometric ratio
(1: 10) of the formulation. In bone [‘53Sm]
concentra
tions are in the range of 20—30times greater than ‘4C.
However, this difference was greatly reduced in tumor
and kidney, and in fact was reversed in blood, where
[‘4C]EDTA% I.D. g' levels were double those of
[‘53Sm].
In general, these data support the concept that
[‘53Sm]EDTAis taken up by kidney and tumor, but
that other forms, possibly colloids, are being deposited
in RES-rich tissues such as liver and bone (marrow).
The optimal tumor indices (% I.D. g' x T.B.) (20)
for low and high specific activity [‘53Sm]EDTAwere 37
(6 hr) and 5 1 (48 hr), respectively, compared with 302
(24 and 48 hr) for [‘53Sm]citratein this tumor model.
Tumor indices for [67Ga] citrate at low and high specific
activities were 16 (24 hr) and 4.5 (48 hr), respectively,
compared with 48.5 (24 hr) for an even lower specific
activity product (22 @molGa kg' body weight) in this
tumor model.
Whole-body clearance studies in Copenhagen
x
Fisher rats bearing transplanted Dunning R3327-H
prostatic tumors confirmed that low specific activity
[‘53Sm]EDTAwas rapidly excreted via the urine, and
that within a few hours after injection, excretion vir
tually ceased. In contrast, excretion of the high specific
activity chelate was progressive throughout the 48-hr
observation period, although the first 6 hr still ac
counted for over half of the injected dose (Table 3).
Total 48-hr excretion was similar (19.2 ±4.4 vs. 23.7
± 0.5)
for the two
formulations.
The
time
course
of
whole-body elimination was not reflective of blood
clearance. The blood clearance profile, when analyzed
by graphic approximation as a triexponential function,
indicated that the slow clearance represented < 15% of
the dose, with a half-time of
1 hr. The intermediate
component represented 15—20%of the dose with an
estimated half-time of 10 mm, and the rapid component
accounted for the remainder of the dose (65—70%)
which was cleared from the blood with a half-time of
1-2 mm. In large part these data represent dilution and
distribution (short T½),urinary excretion (intermediate
T½)
and redistribution (long T½)
processes. Fecal excre
tion, at ‘@‘
1% I.D. per 48 hr, plays a negligible role in
either blood clearance or whole-body elimination de
spite substantial hepatic uptake.
Scintigraphic studies with the low and high specific
activity [‘53Sm]EDTAand [67Ga]citrate formulations
clearly demonstrated the superiority of scintigrams ob
tamed with high specific activity [‘53Sm]EDTAover
scintigrams from other products. Quantitative biodis
tribution measurements
were not made for the rat
model, but it is evident that rapid and almost total
‘“Sm
blood clearance and adequate tumor uptake re
suited in clear delineation ofthe tumor mass from other
soft tissues. Although ‘“Sm
was present in liver and the
Overall, the [‘53Sm]EDTA
tumor uptake (% I.D. g')
skeleton, the [‘53Sm]EDTAimages were superior to
was lower than for other ‘“Sm
chelates ([‘53Sm]HIDA [67Ga]citrate even in these areas because of the absence
5.2—2.7
and 5.2—1.4%
I.D. g'; [‘53Sm]HEDTA
5.3—2% of bloodbackgroundand intestinalradioactivity.
and 3.5—2.
1% I.D. g'; and [‘53Sm]diethylenetriamine
The absence of observable histopathologic effects
pentaacetic acid 3.8—0.3and 2.4—0.4%I.D. g', respec
after relatively large doses of [‘53Sm]EDTA
(up to 10.9
tively, for melanotic and amelanotic B16 melanomas
in C57 black mice) (13) or for the B6D2F, Lewis lung
model after injection of [‘53Sm]citrate(5.8—10.7% I.D.
g@@')
(15). Very low blood levels, however, compensate
in part for this lower uptake when imaging is the final
objective.
Volume30 •
Number2 •
February1989
@imolkg'
vs. 10.9 nmol kg'
for imaging) in prelimi
nary studies, along with the observed biodistribution
and excretion patterns in mice and rats, are supportive
ofthe clinical application of [‘53Sm]EDTA.With good
tumor delineation within 6 hr and a shorter T½com
pared to 67Ga, absorbed radiation doses from this radi
207
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otracer are expected to be 20% higher than for [67Ga]
citrate, which requires a 48—72hr wait after injection
and has a 50% longer physical T½.
Although no attempt
5.
has been made to provide detailed dose calculations,
biodistnbution
data and elimination half-time esti
mates of 57, 34, and 27 hr (liver, bone, and kidney,
respectively) suggest doses of 0.4, 0.09, and 0.3 mGy
MBq' to the respective organs. Recently published data
(21) for [‘53Sm]EDTMP,a bone-seeking radiotherapeu
tic agent, list doses of 0.028, 1.03, and 0.108, mGy
MBq', respectively, for liver, bone marrow and kidney.
In summary, [‘53Sm]EDTA
formulated with a tenfold
excess of Na2EDTA at a final pH of 6.5 is sufficiently
6.
7.
8.
9.
stable for use as a radiotracer in vitro and in vivo. At
10.
lower specific activity ( 14.3 GBq mmol'),
[‘53Sm]
EDTA showed a greater tendency for liver and bone
uptake than at higher specific activity (22. 1 TBq mmol
11.
‘Sm).
With high specific activity ‘53Sm,
it was possible
to reduce by two orders of magnitude (e.g., to 1%) the
mass of Sm injected. This presumably reduced the
12.
formation of colloid in vivo and at the same time
improved uptake (as % I.D. g') by the tumor. Excellent
tumor delineation, low acute and chronic toxicity, and
13.
the potential for “sameday―
dosing and imaging, pro
vide a rationale for further investigation ofhigh specific
activity [‘53Sm]EDTAfor diagnostic tumor scintigra
phy.
14.
15.
ACKNOWLEDGMENTS
16.
JWT was the recipient of an Alberta Heritage Foundation
for Medical Research Graduate Studentship. The authors
17.
thank Ms. Carolyn Johnson for typing the manuscript and
the U. Alberta Slowpoke Facility for production of ‘53Sm2O3.
18.
REFERENCES
1. Johnston GS. Clinical applications of gallium in on
cology.
mt J NuclMed Biol1981;8:249—255.
2. Mather SJ. Labelling with indium-l 11. In:Kristensen
K, Norbygaard E, eds. Saftty and efficacy of radio
pharmaceuticals. Dordrecht: Martinus Nijhoff Pub
ushers, 1987: 5 1—66.
3. O'Mara RE, McAfee JG, and Subramanium G. Rare
earth nuciides as potential agents for skeletal imaging.
JNuclMed 1969;10:49—51.
4. Ketring AR. ‘53Sm-EDTMPand ‘86Re-HEDP
as bone
208
Tse,Wiebe,
andNoujaim
19.
20.
21.
therapeutic radiopharmaceuticals.
Nucl Med Biol
1987; 14:223—232.
Goeckeler WF, Troutner DE, Volkert WA, et al. ‘53Sm
radiotherapeutic bone agents. Nucl Med Biol 1986;
13:479—482.
Goeckeler WF, Edwards B, Volkert WA, et a!. Skeletal
localization of samarium-l53 chelates; potential ther
apeutical bone agents. JNuclMed 1987; 28:495—504.
Lederer CM and Shirley V. Table of isotopes. 7th ed.
New York: John Wiley and Sons, Inc., 1978:858—859.
Hisada K, Ando A. Radioianthanides as promising
tumor scanning agents. J Nucl Med 1973; 14:
615—617.
Higasi T, Ito K, Tobari H, et al. On the accumulation
of rare earth elements in animal tumor. mt j Nuci
MedBiol 1973;1:98—101.
Sullivan JC, Friedman AM, Rayudu GVS, et al. Tu
mor localization studies with radioactive lanthanide
and actinide complexes. mt J Nucl Med Biol 1975;
2:44—45.
Woolfenden JM, Hall JN, Barber HB, et al. [‘535m]
citrate for tumor and abcess localization. mt j Nuci
MedBiol 1983; 10:25 1—256.
Tse JW, Wiebe LI, Noujaim AA. [‘53Sm]Samarium
complexes as radiopharmaceuticals. J Lab Cpd Radi
opharm 1982;19:1548—1549.
Turner JH, Martindale AA, deWitt GB, et al. Samar
ium-l53 chelate localization in malignant melanoma.
EurJNuclMed 1987;13:432—438.
Tse JW, Wiebe LI, Noujaim AA. [‘“Sm]
Samarium
transferrin uptake by experimental tumor models.
Biochem Arch 1988; 4:69—75.
Tse JW, Wiebe LI, Noujaim AA. Comparative studies
of radiotracer citrates in oncological models. 2. ‘53Sm
citrate and 67Ga citrate. Nucl Med Biol: in press.
Coursey BM, Hoppes DD, Schima FJ, et al. The
standardization of samarium-l53. AppI Radial Isot
1987; 38:31—34.
Ehrhardt GJ, Ketring AR, Volkert WA. Production of
Sm-i53 for radiotherapeutic applications. J Lab Cpd
Radiopharm1986;23:1370.
Loevinger R, Budinger TF, Watson EE. MIRD primer
for absorbed dose calculations. New York: The Society
ofNuclear Med, 1987.
Goeckler, WF, Troutner DE, Volkert WA, et al. ‘53Sm
radiotherapeutic bone agents. Nucl Med Biol 1986;
13:479—482.
Emrich D, vz Muehlen A, Wiligeroth F, et al. Com
parison of tumor uptake and kinetics of different
radiopharmaceuticals under experimental conditions.
Ada Radiol(TherPhysBiol).
1972;11:566—572.
Logan KW, Volkert WA, Holmes PA. Radiation dose
calculations in persons receiving injection of samar
ium-153 EDTMP. JNuclMed 1987; 28:505—509.
The Journalof NuclearMedicine